A novel allosteric modulator of the cannabinoid CB1 receptor ameliorates hyperdopaminergia endophenotypes in rodent models

Funding and disclosure The authors declare the following financial and biomedical conflict of interests: Ruth A. Ross, Catharine A. Mielnik, Amy J. Ramsey, Iain R. Greig, Laurent A. Trembleau, Mostafa H. Abdelrahman are co-inventors on a patent application related to ABM300 and structural analogs. Kim S. Sugamori, David B. Finlay, Hayley H.A. Thorpe, Matthieu Schapira, Nirunthan Sivananthan, Chun Kit Li, Vincent M. Lam, Sean Harrington, W. McIntyre Burnham, Jibran Y. Khokhar, Ali Salahpour, Michelle Glass reported no biomedical financial interests or potential conflicts of interest. W. McIntyre Burnham received Δ9- (THC) as a gift from MedReleaf. The authors would like to gratefully acknowledge Wendy Horsfall for mouse colony maintenance. The work was funded by grants to RAR from CIHR (PPP-125784, PP2-139101), CIHR funding to AJR (MOP119298) and CIHR funding to AS (PJT-15619).Peer reviewedPostprintPublisher PD

Neuroarchitecture of Aminergic Systems in the Larval Ventral Ganglion of Drosophila melanogaster

Biogenic amines are important signaling molecules in the central nervous system of both vertebrates and invertebrates. In the fruit fly Drosophila melanogaster, biogenic amines take part in the regulation of various vital physiological processes such as feeding, learning/memory, locomotion, sexual behavior, and sleep/arousal. Consequently, several morphological studies have analyzed the distribution of aminergic neurons in the CNS. Previous descriptions, however, did not determine the exact spatial location of aminergic neurite arborizations within the neuropil. The release sites and pre-/postsynaptic compartments of aminergic neurons also remained largely unidentified. We here used gal4-driven marker gene expression and immunocytochemistry to map presumed serotonergic (5-HT), dopaminergic, and tyraminergic/octopaminergic neurons in the thoracic and abdominal neuromeres of the Drosophila larval ventral ganglion relying on Fasciclin2-immunoreactive tracts as three-dimensional landmarks. With tyrosine hydroxylase- (TH) or tyrosine decarboxylase 2 (TDC2)-specific gal4-drivers, we also analyzed the distribution of ectopically expressed neuronal compartment markers in presumptive dopaminergic TH and tyraminergic/octopaminergic TDC2 neurons, respectively. Our results suggest that thoracic and abdominal 5-HT and TH neurons are exclusively interneurons whereas most TDC2 neurons are efferent. 5-HT and TH neurons are ideally positioned to integrate sensory information and to modulate neuronal transmission within the ventral ganglion, while most TDC2 neurons appear to act peripherally. In contrast to 5-HT neurons, TH and TDC2 neurons each comprise morphologically different neuron subsets with separated in- and output compartments in specific neuropil regions. The three-dimensional mapping of aminergic neurons now facilitates the identification of neuronal network contacts and co-localized signaling molecules, as exemplified for DOPA decarboxylase-synthesizing neurons that co-express crustacean cardioactive peptide and myoinhibiting peptides

The dopamine D1C receptor, expansion and origin of the dopamine D1 receptor family

grantor: University of TorontoDopamine D1 receptors are implicated in several neuronal processes including motivation, reward, cognition and working memory. To date, two D1-like receptors that activate the same second messenger system, the D1/D1A and D5/D1B receptors, have been isolated from mammals. The heterogeneity of dopamine D1 receptor-mediated events, however, may not be completely reconciled by the presence of only two dopamine D1-like receptors, as biochemical, behavioural and pharmacological data have alluded to the existence of other putative subtypes. This work provides direct molecular evidence for the existence of at least three distinct D1-like receptor genes, termed D1A, D1B and D1C, from a number of nonmammalian vertebrate species. These were classified on the basis of phylogenetic, molecular, pharmacological, and functional criteria. While all three subtypes stimulated the ac cumulation of cAMP in expressed cells, D1A, D1B and D1C receptor chimeras in which the highly divergent carboxyl termini were interchanged revealed a modulatory influence of this domain in determining functional responsiveness. The evolutionary emergence and expansion of the D1-like receptor gene family were further investigated from representatives of early diverging phyla. The isolation of an adenylyl-cyclase activating dopamine receptor from Drosophila melanogaster with poor affinity for classical D1-like benzazepine agents indicates that high affinity benzazepine pharmacology arose after the separation of vertebrates and invertebrates. The characterization of a D1-like receptor from amphioxus, a cephalochordate and sister species to vertebrates, indicates that the gene duplication event leading to D1-like diversity occurred with the appearance of early vertebrates. These studies have provided insight into the molecular phylogenetic history and diversity of the dopamine D1-like receptor family. More importantly, the cloning of the D1C receptor lends strong support to the contention that additional D1-like receptor gene products 0may be present in mammals which allow for the expression of the full spectrum of dopamine D1-like receptor mediated events.Ph.D

Delineation of the conserved functional properties of D1A, D1B and D1C dopamine receptor subtypes in vertebrates.

The three main subtypes of dopamine D(1) receptor (D(1A), D(1B) and D(1C)) subtypes found in most vertebrate groups were generated by two major steps of gene duplications, early in evolution. To identify the functional characteristics contributing to conservation of these paralogous D(1) receptors in vertebrates, the pharmacological and functional properties of fish (Anguilla anguilla), amphibian (Xenopus laevis) and human receptors were systematically analysed in transfected cells. The ligand-binding parameters appeared essentially similar for orthologous receptors, but differed significantly among the subtypes. The D(1A) receptors from the three species displayed low intrinsic activity and a fast rate of agonist-induced desensitization. All the orthologous D(1B) receptors exhibited a similar desensitization time-course, but with smaller amplitude of decrease than D(1A) receptors, in agreement with their higher basal activity. In contrast, D(1C) receptors, which do not exist in mammals, have low intrinsic activity and exhibit only weak, but rapid, agonist-induced desensitization, without any changes upon longer treatment with agonist. Thus, each of the three D(1) receptor subtypes are characterized by activation and desensitization properties, in a sequence-specific manner, which has been probably acquired early after gene duplications, and constrained their conservation during vertebrate evolution. These properties have been instrumental to adapt dopamine system to the physiology of the numerous neuronal networks and functions they control in the large and complex brains of vertebrates

Biochemical pathways for the interaction of folate metabolism and the <i>S</i>-adenosylmethionine (SAM) cycle.

<p>Dietary folate is metabolized to 5-methyltetrahydrofolate (5-MTHF), which is used by methionine synthase for the conversion of homocysteine to methionine. Methionine can also be synthesized from betaine, especially in liver tissue. Catabolism of the folates by cleavage of the C9-N10 bond produces <i>p</i>-aminobenzoylglutamate (PABG) and pterin (not shown). Murine Nat2 (and human NAT1) acetylates PABG to <i>N</i>-acetyl-PABG, which is a major folate metabolite found in the urine.</p

2,3,7,8-Tetrachlorodibenzo-p-Dioxin (TCDD)-Inducible Poly-ADP-Ribose Polymerase (TIPARP/PARP7) Catalytic Mutant Mice (TiparpH532A) Exhibit Increased Sensitivity to TCDD-Induced Hepatotoxicity and Lethality

Abstract 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)-inducible poly-adenosine diphosphate (ADP)-ribose polymerase (TIPARP/PARP7), an aryl hydrocarbon receptor (AHR) target gene and mono-ADP-ribosyltransferase, acts as part of a negative feedback loop to repress AHR signaling. This process is prevented by a single H532A mutation in TIPARP that destroys its catalytic activity. We hypothesized that the loss of TIPARP catalytic activity would increase sensitivity to TCDD-induced toxicity in vivo. To test this, we created a catalytically deficient mouse line (TiparpH532A) by introducing a single H532A mutation in TIPARP. Treatment of mouse embryonic fibroblasts or hepatocytes isolated from TiparpH532A mice confirmed the increased TCDD-induced expression of the AHR target genes Cyp1a1, Cyp1b1, and Tiparp. TiparpH532A mice given a single injection of 10 µg/kg TCDD, a nonlethal dose in Tiparp+/+ mice, did not survive beyond day 10. All Tiparp+/+ mice survived the 30-day treatment. TCDD-treated TiparpH532A mice displayed increased expression of AHR target genes, increased steatohepatitis and hepatotoxicity. Hepatic RNA-sequencing revealed 7-fold more differentially expressed genes in TiparpH532A mice than in Tiparp+/+ mice (4542 vs 647 genes) 6 days after TCDD treatment. Differentially expressed genes included genes involved in xenobiotic metabolism, lipid homeostasis and inflammation. Taken together, these data further support TIPARP as a critical negative regulator of AHR activity and show that loss of its catalytic activity is sufficient to increase sensitivity to TCDD-induced steatohepatitis and lethality. Since TIPARP inhibition has recently emerged as a potential anticancer therapy, the impact on AHR signaling, TCDD and polycyclic aromatic hydrocarbon toxicity will need to be carefully considered under conditions of therapeutic TIPARP inhibition

Liver and kidney concentrations of PABG in male and female Nat2−/− C57BL/6J mice.

<p>Results are mean ± SEM, n = 3, and are expressed as percentage of wild-type concentrations.</p